JP2022517578A - Composition for prevention, amelioration or treatment of degenerative brain disease containing pediococcus inopinatus - Google Patents

Abstract

The present invention relates to a composition for preventing, ameliorating or treating a degenerative brain disease containing pediococcus inopinatus as an active ingredient. Since the pediococca suinopinatas strain according to the present invention can not only suppress neuroinflammation but also improve ataxia symptoms in an animal model of Parkinson's disease, it can prevent various degenerative brain diseases including Parkinson's disease. It can be used for improvement and treatment.

Description

The present invention relates to a composition for preventing, ameliorating or treating a degenerative brain disease, which comprises Pediococcus inopinatus, a culture thereof, a lysate thereof, a disrupted product thereof, a fermented product thereof or an extract thereof.

Degenerative brain disease means a disease that occurs in the brain among degenerative diseases that occur with aging. Degenerative brain diseases can be classified according to the major symptoms and the affected brain region, and are represented by Alzheimer's disease and Parkinson's disease. It is known that degenerative brain disease is caused by nerve degeneration due to aging and the aggregation of proteins due to genetic and environmental factors to kill nerve cells, but the exact cause has not yet been clarified.

Parkinson's disease is the second most degenerative brain disease after Alzheimer's disease, and about 1% of the population over the age of 50 suffers from this disease. It has been reported. Parkinson's disease is a disease whose main symptoms are tremor, rigidity, slow behavior, and abnormal posture, and is a chronic disease caused by a deficiency of a neurotransmitter called dopamine in the brain. Dopamine is produced in nerve cells called the substantia nigra of the brain, which are intricately connected to the motor cortex of the brain and various other parts. Parkinson's disease is caused by a deficiency of dopamine, a substance secreted in the substantia nigra to regulate the function of the basal ganglia.

The main symptoms of Parkinson's disease are 1) bradykinesia, 2) resting tremor (tremor-at-rest), 3) muscle rigidity, and 4) loss of postural reflexes. ), 5) flexed posture, 6) freezing phenomenon, etc.

Currently, treatments for Parkinson's disease include drug treatment, surgical treatment, and physical treatment, but in the case of drug treatment, dopamine that is generally deficient in the brain is supplemented, and the neurotransmitter imbalance due to the deficiency of dopamine. Drugs have been used for the purpose of arranging and preventing or delaying the destruction of nerve cells and for controlling other symptoms such as depression. For example, drugs for treating Parkinson's disease include dopamine substitute drugs such as L-DOPA and drugs that act on dopamine receptors, but these drugs are unable to regenerate dead nerve cells. Since it is possible, there is a limit in that it is intended to control symptoms, and long-term administration leads to serious side effects such as involuntary exercise (dyskinesia) and vomiting. Therefore, there is an urgent need to develop a drug that has a neuroprotective effect that ensures safety as well as improving symptoms.

In particular, recently, it has been reported that changes in the intestinal flora induced by patients with Parkinson's disease are pathophysiologically related to the progression of Parkinson's disease, and further, fecal microbiota transplantation (fecal) into an animal model of Parkinson's disease. As a result of microbiota transplantation (FMT), it was known that a neuroprotective effect was exhibited by a neuroinflammation-regulating action (Non-Patent Document 1).

Against this background, research on the prevention and treatment of Parkinson's disease using lactic acid bacteria has been conducted in recent years, but among the symptoms of Parkinson's disease, lactic acid bacteria that directly affect the symptoms of ataxia other than stool Although it has not been clarified, it is desired to develop a lactic acid bacterium that can improve or treat the symptoms of ataxia caused by Parkinson's disease.

(Non-Patent Document 1) Xin Fang, Microbial treatment: the potential application for Parkinson's disease, Neurological Sciences, 2018

The present invention is for preventing, ameliorating or treating a degenerative brain disease containing Pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof as an active ingredient. Attempts to provide the composition.

Therefore, as a result of efforts to find a probiotic strain having a therapeutic effect on degenerative brain disease, the Pediococcus inopinatus strain isolated from kimchi suppresses nerve inflammation. As a result of directly administering the Pediococcus innopinatas strain to a Parkinson's disease mouse model, it was confirmed that the ataxia symptoms of the mice were improved, and the present invention was completed.

The Pediococcus inopinatus strain according to the present invention is preferably a Pediococcus inopinatus strain derived from kimchi, and more preferably a Pediococcus inopinatus strain WIKIM27 (Pediococcus inopinatus WIKI27). Yes, most preferably the Pediococcus innopinatas WIKIM27 strain deposited under accession number KCCM12653P, but is not limited thereto.

The Pediococcus inopinatus strain of the present invention is a Gram-positive bacterium, is facultative anaerobe capable of growing under both aerobic and anaerobic conditions, and produces spores. It does not form, is not motile, and the cells are in the form of cocci.

The Pediococcus inopinatus strain of the present invention is a probiotic strain and has a general intestinal regulating effect and an immune enhancing effect of lactic acid bacteria. It is a well-known fact that lactic acid bacteria of the genus Pediococcus have an intestinal regulating effect and an immune enhancing effect.

In the present invention,'probiotics' means'living microorganisms' that improve the intestinal microbial environment of the host in the gastrointestinal tract of animals including humans and have a beneficial effect on the health of the host. Understood. Probiotics are living microorganisms with probiotic activity that, when fed to humans or animals in dry cell or fermented form in single or complex strain form, are the host gut flora. Can have a beneficial effect on.

One embodiment of the present invention provides a composition comprising a pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof.

The pediococcus inopinatus contained in the composition according to the present invention may be present as a live or dead cell, or may be present in a dried or lyophilized form. Suitable lactic acid bacterium forms and methods of formulation to be included in various compositions are well known to those of skill in the art. For example, pediococca suinopinatas is a culture obtained by culturing in a known liquid medium or solid medium, or a fermented product obtained by culturing the strain and additional components together, or the strain. Can be formulated in the form of an extract extracted with an organic solvent, a lysate (or crushed product) obtained by dissolving the cell membrane of the strain, or being crushed or homogenized, but the present invention is not limited thereto.

In one embodiment, the composition may be a composition comprising a strain of Pediococcus inopinatus WIKIM27 (accession number KCCM12653P).

In another embodiment, the composition may be a composition comprising a Pediococcus innopinatas WIKIM27 strain present as a live or dead bacterium.

In yet another embodiment, the composition may be a composition comprising a culture, lysate, fermented or extract of Pediococcus innopinatas WIKIM27 strain.

The present invention provides an intestinal regulating composition comprising pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof. The intestinal regulating composition according to the present invention can be used for the prevention, treatment and amelioration of gastrointestinal diseases of animals including humans, preferably the animals include domestic animals such as cattle, horses and pigs. The'gastrointestinal disease' includes all gastrointestinal harmful bacterial infections and inflammatory bowel disease, for example, infectious diarrhea due to pathogenic microorganisms (Escherichia coli, salmonella, clostridium, etc.), gastrointestinal inflammation, inflammatory bowel disease, etc. Includes, but is not limited to, neurogenic enteritis syndrome, small bowel microbial hypergrowth, intestinal feeding diarrhea, and the like.

The intestinal regulating composition according to the present invention is preferably administered orally. The dosage may vary depending on the type of gastrointestinal disease, degree of disease, age, gender, race, treatment or preventive purpose, etc., but generally, 10 to 100 billion animals per day based on adults. Can be administered.

The present invention provides an immunopotentiating composition comprising a pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof. It is a well-known fact that lactic acid bacteria of the genus Pediococcus have an intestinal regulating effect and an immune enhancing effect.

The present invention also contains pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof as an active ingredient, and improves memory, cognitive ability, muscle strength or exercise performance. Provided is a food composition for capacity improvement.

In the food composition, Pediococcus inopinatus can be applied as it is with all the above-mentioned contents.

In one embodiment, the composition may be a composition comprising a strain of Pediococcus inopinatus WIKIM27 (accession number KCCM12653P).

In another embodiment, the composition may be a composition comprising a Pediococcus innopinatas WIKIM27 strain present as a live or dead bacterium.

In yet another embodiment, the composition may be a composition comprising a culture, lysate, disrupted product, fermented product or extract of Pediococcus innopinatas WIKIM27 strain.

In the present invention, "improvement of memory" and "improvement of cognitive ability" refer to memory deterioration, memory impairment, or memory impairment that occurs when brain atrophy and destruction of brain nerve cells occur due to physical fatigue, lack of sleep, excessive alcohol intake, dementia, and the like. Decreased cognitive ability means the effect of regulating harmful substances that damage brain cells to maintain cognitive ability, or regulating neurotransmitters in the brain to improve the decreased cognitive ability. The memory ability is the ability to receive necessary information, store it in the brain, and take it out and use it when necessary, and the cognitive ability is the ability to discriminate and recognize things.

In the present invention, "improvement of muscle strength" and "improvement of exercise performance" regulate energy metabolism that the exercise performance is reduced due to various diseases, hormonal changes due to age, obesity, liquor, tobacco and the like. It means the effect of improving exercise performance. The muscle strength is the force that a muscle can exert to a certain resistance (weight, force) at one time, and is the ability of the muscle, that is, the maximum force that a muscle or muscle tissue can exert at one time. Exercise performance refers to the ability to exercise using such muscle strength.

When the composition of the present invention is used as a food composition, the food composition may include a health functional food or a form such as a seasoning, a beverage, or a bar. In addition, the food composition containing the strain as an active ingredient can include a beverage such as fermented milk. Therefore, the present invention provides a lactic acid bacterium starter for fermentation, which comprises Pediococcus inopinatus or a culture thereof.

In addition to the active ingredient, the food composition of the present invention may be produced using a food-appropriate and physiologically acceptable auxiliary agent, and the auxiliary agent may be an excipient. Disintegrants, sweeteners, binders, coatings, swelling agents, lubricants, lubricants or flavoring agents can be used.

The food composition may be preferably formulated as a pharmaceutical composition by including one or more pharmaceutically acceptable carriers in addition to the above-mentioned active ingredient for administration.

For example, for formulation into the form of tablets or capsules, the active ingredient may be combined with an oral, non-toxic, pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like. Appropriate binders, lubricants, disintegrants and color formers may also be included as mixtures, if desired or required. Suitable binders are, but are not limited to, natural sugars such as starch, gelatin, glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacant or sodium oleate, sodium stearate, Includes magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, etc. Disintegrants include, but are not limited to, starch, methylcellulose, agar, bentonite, xanthan gum and the like. Acceptable pharmaceutical carriers in compositions formulated as liquid solutions include saline, sterile water, ringer solutions, buffered saline, albumin injection solutions, dextrose solutions, which are suitable for sterilization and living organisms. Maldextrin solution, glycerol, ethanol and one or more of these components can be mixed and used, and if necessary, other usual additives such as antioxidants, buffers and bacteriostatic agents can be added. May be good. Further, a diluent, a dispersant, a surfactant, a binder and a lubricant are further added to form an injectable dosage form such as an aqueous solution, a suspension, an emulsion, a pill, a capsule, a granule or a tablet. May be.

The food composition according to the present invention can be added to various foods. Foods to which the composition of the present invention can be added include, for example, beverages, vitamin complex agents, dietary supplements and the like.

The food composition of the present invention may contain ingredients normally added during food production, including, for example, proteins, carbohydrates, fats, nutrients, seasonings and flavoring agents. Examples of carbohydrates mentioned above include monosaccharides such as glucose, fructose; dissaccharides such as maltose, sucrose, oligosaccharides; and polysaccharides such as dextrin, cyclodextrin and the like and common sugars and erythritol. It is a sugar alcohol such as sorbitol and erythritol. As the flavoring agent, a natural flavoring agent [taumatin, stevia extract (for example, levaugioside A, glycyrrhizin, etc.]) and a synthetic flavoring agent (saccharin, aspartame, etc.) can be used. For example, when the food composition of the present invention is produced as a drink and a beverage, it may further contain citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, various plant extracts and the like. can.

The present invention also provides a pharmaceutical composition for preventing or treating a degenerative brain disease containing pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof as an active ingredient. offer.

In one embodiment, the composition may be a composition comprising a strain of Pediococcus inopinatus WIKIM27 (accession number KCCM12653P).

In another embodiment, the composition may be a composition comprising a Pediococcus innopinatas WIKIM27 strain present as a live or dead bacterium.

In yet another embodiment, the composition may be a composition comprising a culture, lysate, disrupted product, fermented product or extract of Pediococcus innopinatas WIKIM27 strain.

In addition, the present invention administers a therapeutically effective amount of pediococcus inopinatus, its culture, its lysate, its crushed product, its fermented product or its extract to a subject in need thereof. Provided a method for treating degenerative brain diseases including.

The term "object" as used herein refers to a mammal that is the subject of treatment, observation or experiment, and may preferably be a human or animal in need of prevention and / or treatment of degenerative brain disease. ..

The term "degenerative brain disease" as used herein refers to Parkinson's disease, Alzheimer's disease, Huntington's disease, Lugeric disease, and amyotropic disease. Creutzgeldt-Jacob disease, stroke, multiple sclerosis, neuroinflammation, learning disorder and cognitive impairment from cognitive impairment to cognitive impairment. It may be, but is not limited to, one or more diseases of choice.

In one embodiment, the degenerative brain disease may be Parkinson's disease.

In the examples below, it was confirmed that treatment with the Pediococcus innopinatas strain suppressed neuroinflammation. In addition, as a result of rotarod evaluation and grip strength evaluation after direct administration of Pediococcus inopinatus strain to a Parkinson's disease mouse model, the balance retention ability on the rotating device was higher than that of the negative control group (PBS intake group). It was confirmed that the grip strength was high. That is, the present invention can be used for the treatment of degenerative brain diseases including Parkinson's disease by confirming that the pediococca suinopinatas strain is used to improve the ataxia symptom of the Parkinson's disease mouse model. It was confirmed.

Neuroinflammation is Alzheimer's disease, Parkinson's disease, amyotropic disease, multiple sclerosis, multiple sclerosis, multiple sclerosis, etc. It causes. The association of neuroinflammation with the pathogenesis of degenerative brain disease has been described in several literatures (Mol Med Rep. 2016 Apr, 13 (4): 3391-3396; J Immunol Res. 2018 Apr16, 2018: 4784268; Transl Neurologicer. Well-known in 2015, 4:19, etc.), the technique of using a neuroinflammation regulator for the treatment of degenerative brain disease also falls under the known technique.

The pharmaceutical composition according to the present invention can be administered by a conventional method through a route such as intravenously, intraarterial, intraperitoneal, intramuscular, or intrasternal.

The pharmaceutical composition of the present invention can include a pharmaceutically acceptable carrier. In the present invention, the term "pharmaceutically acceptable carrier" means a carrier or diluent that does not significantly irritate an organism and does not interfere with the biological activity and properties of the components administered. The pharmaceutically acceptable carrier in the present invention is a mixture of one or more components of saline, sterile water, ringer solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol and their components. It can be used and, if desired, other conventional additives such as antioxidants, buffers and bacteriostatic agents can be added and formulated in the form of injections suitable for injection into tissues or organs. Further, it may be formulated as an isotonic sterilized solution or, if necessary, a dry preparation (particularly, a lyophilized preparation) which can be injected by adding sterilized water or physiological saline. Further, a target organ-specific antibody or other ligand may be used in combination with the carrier so as to be able to act specifically on the target organ. Suitable formulations known in the art can be those disclosed in the literature (Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA).

Further, preferably, the composition of the present invention can further contain a filler, an excipient, a disintegrant, a binder, a lubricant and the like. The compositions of the invention can also be formulated using methods known in the art such that they can be administered to mammals to provide rapid, sustained or delayed release of the active ingredient.

In the present invention, "administration" means to induce a patient to introduce the composition of the present invention by an appropriate method, and the route of administration of the composition of the present invention is oral or non-oral as long as it can reach the target tissue. It can be administered by various oral routes. Intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intrablood administration, oral administration, local administration, intranasal administration, intrapulmonary administration, and rectal administration are possible, but not limited to this.

For example, the compositions of the present invention can be administered by intramuscular vein or intraperitoneal injection at the time of clinical administration.

For injection, it may preferably be formulated as a pharmacologically compatible buffer such as Hank's solution, Ringer's solution, or saline buffer. For mucosal permeation administration, a non-penetrating agent suitable for the passing barrier is used in the dosage form. These non-penetrating agents are generally known in the art.

Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions and the like. As the non-aqueous solvent and the suspending solvent, propylene glycol, polyethylene glycol, a vegetable oil such as olive oil, an injectable ester such as ethyl oleate, and the like may be used.

As used herein, the term "effective amount" means an amount necessary to delay or completely stop the onset or progression of a specific disease to be treated, and the pharmaceutical composition of the present invention. The effective amount of pediococca suinopinatas WIKIM27 contained in the above means an amount required for obtaining a preventive or therapeutic effect on degenerative brain disease. Therefore, the effective amount is the type of disease, the severity of the disease, the type and content of other components contained in the composition, and the patient's age, weight, general health condition, sex and diet, administration time, It may be regulated by a variety of factors, including the route of administration, duration of treatment, and co-use drugs. It will be apparent to those skilled in the art that the appropriate total daily dose may be determined by the treating physician within accurate medical judgment.

For the purposes of the present invention, the specific therapeutically effective amount for a specific patient is the type and degree of the reaction to be achieved, and in some cases, the specific composition including the presence or absence of the use of other preparations, the age of the patient. , Body weight, general health, gender and diet, administration time, route of administration and composition rate, duration of treatment, various factors and pharmaceuticals including drugs used with or simultaneously with the specific composition. It is preferred to apply individually by similar factors well known in the field.

As used herein, "treatment" means approach to obtain beneficial or suitable clinical results. For the purposes of the present invention, beneficial or suitable clinical results, whether detectable or undetectable, are non-limiting, alleviating symptoms, reducing the extent of the disease, resting the disease state (ie, resting the disease state). Does not worsen), delays or slows disease progression, improves or temporarily alleviates disease status, and includes remission (partially or wholly). “Treatment” may also mean prolonging survival compared to the expected survival rate without treatment. "Treatment" means all therapeutic treatments and prophylactic or preventative measures. These treatments include those required for disorders that have already occurred, as well as those that are prevented. "Alleviating" the disease reduces or / or delays or prolongs the time course of progression (time course) compared to no treatment. Means to be done.

The present invention also provides a feed additive or feed containing pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof as an active ingredient.

When used as a feed additive, the composition may be a 20-90% highly concentrated solution or may be produced in powder or granular form. The feed additive may be an organic acid such as citric acid, fumaric acid, adipic acid, lactic acid or malic acid, or phosphorus such as sodium phosphate, potassium phosphate, acidic pyrophosphate or polyphosphate (polymerized phosphate). One or more of acid salts or natural antioxidants such as polyphenols, catechins, alpha-tocopherols, rosemary extracts, vitamin C, green tea extracts, licorice extracts, chitosan, tannic acid, phytic acid, etc. Can be further included. When used as a feed, the composition can be formulated in a normal feed form and can also contain normal feed components.

The feed additives and feeds are grains such as crushed or crushed wheat, swallow, wheat, corn and rice; vegetable protein feeds such as oil vegetables, beans and sunflower-based feeds; animal protein feeds. , For example, blood meal, meat meal, bone meal and fish meal; sugar and dairy products, for example, a dry component consisting of various milk powder and milk meal powder, etc. may be further contained, and other nutritional supplements, digestion and absorption enhancers, etc. Further can include growth promoters and the like.

The feed additive can be administered to the animal alone or in combination with other feed additives in an edible carrier. In addition, the feed additive can be easily administered to animals as a top dressing, by direct mixing with animal feed, or as an oral dosage form separate from feed. When the feed additive is administered separately from the animal feed, as is well known in the art, in combination with a pharmaceutically acceptable edible carrier, as an immediate release or sustained release dosage form. Can be manufactured. Such edible carriers may be solid or liquid, such as corn starch, lactose, sucrose, bean flakes, peanut oil, olive oil, sesame oil and propylene glycol. When solid carriers are used, the feed additive may be tablets, capsules, powders, troches or sugar-containing tablets or top dressings in undispersed form. When liquid carriers are used, the feed additives may be in the form of gelatin soft capsules, syrups, suspensions, emulsions, or solutions.

In addition, the feed additive and feed may contain an auxiliary agent, for example, a preservative, a stabilizer, a wetting agent, an emulsifier, a solution accelerator, or the like. The feed additive can be drunk, sprayed or mixed and added to animal feed for use.

The feed or feed additive of the present invention can be applied to various animal diets including mammals, poultry and fish.

As the mammal, in addition to pigs, cows, sheep, goats, experimental rodents, and experimental rodents, pet animals (for example, dogs and cats) can also be used, and as the poultry, chickens, It can also be used for schimen butterflies, ducks, geese, pheasants, quails, etc., and can be used for trout as the fish, but is not limited thereto.

The feed or feed additive of the present invention is applied to an animal diet and may be used for animal growth, immunity enhancement, and the like.

In addition, the feed or feed additive of the present invention can be used for improving memory, cognitive ability, muscle strength, exercise performance, and prevention, improvement, or treatment of degenerative brain diseases in animals.

The amount of the pediococcus inopinatus strain contained in the composition according to the present invention may be about 10 ⁶ to 10 ¹² cfu / ml based on one time, for example, 10 ⁷ to 10 ¹¹ cfu / ml, 10 ⁸ It may be ~ 10 ¹⁰ cfu / ml. When the strain is administered, it is preferable to administer it in a viable state, and it can be administered in a dead or attenuated state before ingestion. Further, in the case of producing using a culture supernatant or the like, a sterility process by a heat treatment process can be further included. The amount of strain required to have the minimum efficacy and the degree of daily intake may vary depending on the physical condition or health condition of the ingestor, but generally, about ¹⁰⁶ to 10 ¹² cfu / ml may be used, for example, 10 ⁷ to. It may be 10 ¹¹ cfu / ml, 10 ⁸ to 10 ¹⁰ cfu / ml.

The advantages and features of the invention, and how to achieve them, will be clarified in detail with reference to the examples below. However, the present invention is not limited to the examples disclosed below, and may be embodied in various other forms. It is provided to fully inform a person having ordinary knowledge in the field of the technology to which the invention belongs, and the present invention is defined only by the claims.

Since the pediococca suinopinatas strain according to the present invention can improve not only neuroinflammation inhibitory effect but also ataxia symptoms in Parkinson's disease animal model, prevention of various degenerative brain diseases including Parkinson's disease, It can be used for improvement and treatment.

It is a figure which shows the experimental process for confirming the neuroinflammatory inhibitory effect of the pediococcus inopinatas WIKIM27 strain of this invention. The results of treatment of the glial cells inflamed by LPS with the culture medium of the pediococcus inopinatus WIKIM27 strain of the present invention were observed by immunofluorescent staining. It is a graph which shows the result of treating the culture solution of the pediococcus inopinatas WIKIM27 strain of this invention on the glial cell inflamed by LPS. This is the result of oral administration of the Pediococcus innopinatas WIKIM27 strain of the present invention to a Parkinson's disease mouse model, and then evaluation of the motor ability of the mice using a Rotarod. This is the result of evaluating the grip strength of mice after oral administration of the Pediococcus innopinatas WIKIM27 strain of the present invention to a Parkinson's disease mouse model.

Hereinafter, the present invention will be described in detail with reference to examples. The following examples are for exemplifying the present invention, and the scope of the present invention is not limited to the following examples.

[Example]
Example 1: Preparation of Pediococcus inopinatus strain The Pediococcus inopinatus WIKIM27 strain, which was derived from kimchi and was entrusted to the Korea Microbial Conservation Center under the accession number KCCM12653P, was used in the experiment. The pediococcus innopinatas WIKIM27 strain was cultured in MRS medium at 30 ° C. for 24 hours, and then the cells were centrifuged at 8,000 rpm for 5 minutes and washed with PBS to remove the remaining medium components. This is further inoculated into DMEM (Dulvecco's Modified Eagle's Medium, Hyclone, USA) medium at a bacterial count of ¹ × 109 CFU / mL, cultured at 30 ° C. for 24 hours, and then centrifuged at 8,000 rpm for 5 minutes. Then, the cells were removed, the pH of the supernatant was corrected to 7.2, and then filtered using a syringe filter (0.22νm pore size) before use.

Example 2: Primary culture of glial cells The scalp of a 2-day-old Sprague-Dawley rat was disinfected with alcohol, and then the cerebrum was rapidly removed without damage. The removed cerebrum was immersed in a cold HBSS (Hank's Balanced Salt Solution) solution, and only the cerebral cortex was separated under an anatomical microscope. .. For culture of glial cells, 10% fetal bovine serum (Hyclone, USA), 10% horse serum (horse serum, Hyclone, USA) and 1% penicillin / streptomycin (Gibco, USA). ) Was used, and the cells were cultured in an incubator containing 5% CO ₂ under 37 ° C. conditions.

Example 3: Confirmation of neuroinflammation-suppressing effect of Pediococcus inopinatus strain In order to confirm the neuroinflammation-suppressing effect of Pediococcus inopinatus WIKIM27 strain prepared in Example 1, LPS (lipopolysaccharide) was used to confirm the neuroinflammation-suppressing effect. After inducing cell inflammation, it was treated with WIKIM27 culture medium. After dispensing the glial cells into a 24-well plate at a concentration of 1.0 × 10 ⁵ cells / ml, the cells were replaced with serum-free medium 24 hours later, and LPS was concentrated at 100 ng / ml 16 hours later. After 2 hours of LPS treatment, the WIKIM27 culture medium was treated so as to have a volume of 1/5 of that of the glial cell medium. The experimental process of this experiment is schematized in FIG. At this time, as a control group, a Weissella cybaria WIKIM28 (Weissella cybaria WIKIM28; accession number KFCC11625P) strain and a Lactobacillus sakei WIKIM30 (Lactobacillus sakei WIKIM30; accession number KFCC11618P) strain were used.

As shown in FIGS. 2 and 3, in the case of the WIKIM27 treatment group (W27_CM), the activity of glial cells (microglial cells and astrocytes) is significantly reduced even though LPS induces neuroinflammation. Was confirmed by immunostaining chemistry. At this time, Iba-1 (ionized calcium-bound adhesive protein 1) and GFAP (Glial fiberly acid protein) were used for the microglial cells and the astrocytes, respectively, as selective markers. That is, from the above results, it was confirmed that the WIKIM27 strain culture solution alleviated the neuroinflammation.

Example 4: Preparation and administration of Pediococcus innopinatas strain The Pediococcus innopinatas WIKIM27 strain was cultured in MRS medium at 30 ° C. for 24 hours, and then the cells were centrifuged at 8,000 rpm for 5 minutes. It was washed with PBS to remove the remaining medium components. The number of bacteria was quantified to 1 × 10 ¹⁰ CFU / mL using PBS, and 0.2 ml (1 × 10 ⁹ CFU) was orally administered to the experimental animals 5 times a week using a sonde, and sterilized in the negative and positive control groups. PBS was administered.

Example 5: Confirmation of preventive and therapeutic efficacy of Parkinson's disease by treatment of Pediococca suinopinatas strain in an animal model To confirm the therapeutic efficacy of the Pediococca suinopinatas WIKIM27 strain prepared in Example 1 above for Parkinson's disease. , Parkinson's disease mouse model was subjected to rotarod evaluation (rota-rod test) and grip strength evaluation (grip strength test). At this time, in the Parkinson's disease mouse model, 8-week-old male mice (C57BL / 6) were adapted to the breeding room for 1 week, and after oral administration of Pediococca suinopinatas WIKIM27 strain for 30 days, MPTP (1-methyl) was used. Parkinson's disease was induced by intraperitoneal injection (30 mg / kg once daily for 5 days) of -4-phenyl-1,2,3,6-tetrahydropridine). In the following experiments, Naive shows an untreated group, MPTP shows a negative control group, selegiline shows a positive control group, and selegiline is a neuroprotective MAO-B inhibitor and is MPTP-injected. Oral administration (3 mg / kg) was performed 3 days before.

1) Rotor rod evaluation The rotor rod evaluation was performed by measuring the period during which the mouse model was balanced on the rotating drum on the rotor rod composed of the base platform and the rotating rod having a non-smooth surface. To adapt the mouse model to the device, the mouse model was trained at a rate of 4 rpm for 2 minutes and then evaluated 30 minutes later. The evaluation applied an acceleration paradigm of 1 rpm per 8 seconds from a speed of 4 rpm to a maximum speed of 40 rpm, after which the rotational speed was kept constant at 40 rpm for a maximum of 300 seconds.

As a result (Fig. 4), in the negative control group treated with PBS, the equilibrium was maintained up to an average of 7.7 rpm, and in the experimental group to which the Pediococcus innopinatas WIKIM27 strain was orally administered, the average was up to 10.6 rpm. It was confirmed that the balance was maintained.

2) Grip strength evaluation The grip strength evaluation was performed by measuring the grip strength of the forefoot of the mouse model with a grip strength meter. The grip strength of the forefoot of the mouse model was measured 5 times each for a total of 3 times, and the grip strength of the mouse model was displayed by the average value of the 3 highest numerical values.

As a result (Fig. 5), the grip strength of the forefoot of the negative control group was 77.78 gram, whereas that of the experimental group ingesting the Pediococcus inopinatus WIKIM27 strain was 85.79 gram, which is much higher than that of the negative control group. It was confirmed that it has a high grip strength.

That is, by feeding the Parkinson's disease mouse model with the Pediococca suinopinatas WIKIM27 strain, both the rotarod and the grip force evaluation showed higher results than the negative control group, and the ataxia of the Parkinson's disease mouse model was shown. It was confirmed that the symptom was significantly improved (P <0.05).

KCCM12653P

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Claims (12)

A food composition for improving memory, cognitive ability, muscle strength or exercise performance, which comprises Pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof or an extract thereof as an active ingredient. .. The food composition according to claim 1, wherein the food is a health functional food. The food composition according to claim 1, wherein the food is a beverage, a bar or fermented milk. The food composition according to claim 1, wherein the pediococcus inopinatas is pediococcus inopinatas WIKIM27 deposited under the accession number KCCM12653P. A pharmaceutical composition for the prevention or treatment of degenerative brain disease, which comprises Pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof as an active ingredient. The degenerative brain diseases include Parkinson's disease, Alzheimer's disease, Huntington's disease, Lugeric's disease, and amyotropic lattice. One selected from the group consisting of disease, stroke, multiple sclerosis, neuroinflammation, learning disorder, cognitive impairment, and memory impairment. , The pharmaceutical composition according to claim 5. Administered by the route of administration selected from the group consisting of oral administration, intravenous administration, intraperitoneal administration, intramuscular administration, subcutaneous administration, blood administration, local administration, intranasal administration, intrapulmonary administration and rectal administration. Item 5. The pharmaceutical composition according to Item 5. The pharmaceutical composition according to claim 5, wherein the pediococcus innopinatas is contained in an amount of 1 × 10 ⁶ CFU / ml to 1 × 10 ¹² CFU / ml. The pharmaceutical composition according to claim 5, which prevents or treats ataxia symptoms in an individual having a degenerative brain disease. The pharmaceutical composition according to claim 5, wherein the pediococcus inopinatas is pediococcus inopinatas WIKIM27 deposited under the accession number KCCM12653P. Degenerative brain disease, including administration of a therapeutically effective amount of pediococcus inopinatus, its culture, its lysate, its lysate, its fermented product or its extract to a subject in need thereof. Treatment method. A feed additive comprising Pediococcus inopinatus, a culture thereof, a lysate thereof, a crushed product thereof, a fermented product thereof or an extract thereof.

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